S165: INTEGRATED GENETIC DIAGNOSTICS OF PATIENTS WITH EARLY ONSET OF DE NOVO MYELODYSPLASTIC SYNDROMES.
Authors: E. Attardi, L. Tiberi, D. Formicola, R. Artuso, V. Santini
Journal: HemaSphere (2022)

Background: Cytogenetic and molecular alterations determine myelodysplastic syndrome (MDS) prognosis and must be evaluated during disease history. Relevance of germline (GM) predisposition in MDS was stressed in WHO classification, but its actual incidence is probably underestimated. Recently, the widespread availability of large-scale genomic sequencing techniques has facilitated the investigation of GM variants, and the identification of novel GM variants predisposing to MDS is expected. In de novo MDS patients (pts) with an early onset of the disease (age ≤55 yrs) when no syndromic signs drive the genetic assessment, whole exome sequencing (WES) analysis of bone marrow (BM) and saliva can constitute an optimal diagnostic procedure, improving the performance of targeted next-generation sequencing (t-NGS) approach. Aims: We adopted integrated diagnostics to define the cytogenetic profile, and WES to investigate possible GM predisposition of a cohort of 30 de novo MDS pts with an unusual young age at disease onset (≤55 yrs). Standard karyotype (SK) was paralleled by low coverage whole genome sequencing (lc-WGS) on plasma cell-free DNA (cf-DNA), as non-invasive screening test for detection of structural chromosome abnormalities. Methods: Patient BM-DNA / peripheral blood DNA (PB-DNA) was analysed by WES and compared to DNA extracted from saliva (S-DNA) samples as control for GM variants. To identify high-confidence MDS variants, we attributed an internal deleteriousness score to define variants, from “of uncertain significance” to “pathogenic” (ACMG guidelines). Variants were suspected as GM where variant allele frequency was > 0.3 and had been confirmed for genes detected in both S-DNA and BM/PB-DNA. To confirm the somatic nature of some variants, deep t-NGS of 27 myeloid custom panel gene was applied in 26/30 MDS cases on BM-DNA. SK and lc-WGS (0.2x) on plasma cf-DNA were assessed in 27/30 pts. Results: The small cohort of “young” MDS pts here analysed (Table 1). The majority of pts, 17/30 (57%) presented at least one high-confidence variant evaluated as GM for MDS. A total of 26 GM variants were recognized. Patients could be grouped on the basis of the type of variant as follows: 11/30 pts (37%) presented variants in genes involved in DNA repair defects and cancer predisposition (ATM, ATR, FANCA, FANCM, PARN, BRCA1, BRCA2, CHEK2); 4/30 (13%) had variants involved in genetic predisposition to myeloid neoplasm - GATA2, ANKRD26 and RBBP6 - the latter classically associated to MPN familial cases; one presented compound heterozygous variants in SBDS; 4 presented variants related to hereditary red blood cell defects. In our cohort, we identified cases with a complex mode of inheritance (4/17) as well as high confidence GM variants in 4 pts in 2 new interesting cancer predisposition genes: RBBP6 and PARN. Lc-WGS of plasma cf-DNA confirmed all the cytogenetic alterations found by SK (8/27) and identified a 21q22.12 deletion involving RUNX1 locus in a patient, not revealed by SK. Image:Summary/Conclusion: By means of an integrated diagnostics, defined as the convergence of diagnostic techniques with advanced information technology (WES, t-NGS, lc-WGS), as a proof of principle, it was possible to fully characterize the genetic asset of a particular subgroup of 30 MDS cases. We also demonstrated that lc-WGS of plasma cf-DNA has an excellent sensitivity allowing to perform periodic cytogenetic evaluations in MDS pts without invasive maneuvers. We showed that, by the use of WES, GM alterations were demonstrated in a small cohort of “young” non-syndromic MDS pts in 57% of cases, a proportion unexpectedly high.